Wednesday 10 July 2013, Strathblane & Cromdale Halls, 16:30-18:30
Poster session B - Membranes
P.154 The influence of salt on zwitterionic lipid bilayers
R Barker1 and S Roser2
1
Institut Laue-Langevin, France, 2University of Bath, UK
Calcium ions play a central role in a number of physiological processes, both indirectly as a messenger spreading
excitation between an array of receptors leading to gene activation and directly involved in specific ion channel
membrane depolarisation[1]. Similarly, cellular fusion involves a complex interplay of proteins[2] and protein
receptors[3] but this is all thought to be triggered and regulated by the concentration gradient of Ca2+ ions
present[4]. Often the role of salt in protein interactions with lipid bilayers is neglected. This work presents a detailed
neutron reflectivity study of both monolayer and floating lipid bilayer systems to understand the extent to which
CaCl2 influences the membrane of zwitterionic lipid systems as a function of concentration and temperature.
Six distinct regimes have been identified in the mechanism of membrane change as the CaCl 2 concentration is
increased in the bulk solution. These can be explained by considering the contributions of steric, electrostatic and
Helfrich repulsions to the overall three dimensional bilayer structure. The data is presented within the wider context
of a number of biological examples, where the conclusions of this study can be used to more efficiently design
membrane studies using neutron reflectivity in the future.
[1]
[2]
[3]
[4]
P. Racay & J. Lehotcky, Gen. Physiol. Biophys., 15, 273 (1996)
T.-Y. Yoon, X. Lu, J. Diao, S.-M. Lee, T. Ha & Y.-K. Shin, Nat. Struct. Mol. Biol., 15, 707 (2008)
T. Sollner, S.W. Whiteheart, M. Brunner, H. Erdjument-Bromage, S. Geromanos, P.L. Tempst & J. Rothman,
Nature, 362, 318 (1993)
R. Jahn & T.C. Sudhof, Annu. Rev. Biochem., 68, 864 (1999)
P.155 Effects of phase separation and gravity on the interfacial properites resulting from PAMAM dendrimer/lipid
vesicle interactions
R Campbell1, A Åkesson2 and M Cárdenas2
1
Institut Laue-Langevin, France, 2University of Copenhagen, Denmark
Supported Lipid Bilayers (SLBs) are important substrates for the study of drug/polymer/nanoparticle interactions
and they are commonly employed in studies using non-invasive, surface-sensitive techniques such as neutron
reflectometry (NR). We have investigated the interactions of PAMAM dendrimers with lipid vesicles and SLBs using
NR and have found contradictory results depending on the location/geometry of the interface (i.e. vertical vs
horizontal up and down configurations) – which itself is usually imposed by the choice of instrument used. Recently
we have carried out work on FIGARO at the ILL involving a novel comparison of reflection up vs down measurements,
which demonstrated that phase separation and gravity determine the interfacial properties according to whether the
interface is located below or above the bulk liquid. Very different surface nanostructures emerged at the different
surfaces of the same sample, which tells us that an equilibrium model with the solution as a single phase giving a
unique answer cannot be used to predict the surface behaviour unambiguously. Issues concerning (1) the role of
bound vesicles, (2) the influence of the substrate charge and (3) the underlying mechanism of surface
nanostructure formation will be discussed. This work shows that a better understanding of non-equilibrium effects is
required in order to interpret experimental results from complex mixed systems in soft matter and biophysics which
exhibit phase separation.
ICNS 2013 International Conference on Neutron Scattering
P.156 Magnetic response of functionalised lipid bilayers
J Kohlbrecher2, M Liebi1, P Fischer1 and P Walde1
1
ETH Zurich, Switzerland, 2Paul Scherrer Institut, Switzerland
Magnetic field effects on biological molecules are rarely observed, due to the small diamagnetic susceptibility of
individual molecules. In self-assembly systems, such as phospholipid bilayers, where the molecules are aggregated
parallel to each other, the anisotropy of the diamagnetic susceptibility become additive and a magnetic orientation
in a magnetic field becomes possible. A well known system, where this magnetic orientation has been exploited, is
a bicellar mixture of long and short chain phospholipids used in NMR studies of transmembrane proteins. By doping
these aggregates with paramagnetic lanthanides conferring a large magnetic moment, the responsiveness to
magnetic fields can be enhanced. The effect of magnetic fields on phospholipid vesicles and bicelles containing a
chelator-lipid with complexed paramagnetic lanthanides will be shown.
P.157 Effect of functionalized gold nanoparticles on floating lipid bilayers
M Maccarini1, S Tatur2, R Barker3 and G Fragneto3
1
CEA, France, 2University of Illinois, USA, 3Institut Laue Langevin, France
The development of novel nano-engineered materials poses important questions on how these new materials will
interact with living systems. On the one hand, possible adverse effects must be assessed in order to prevent risks for
health and environment [1]. On the other hand, the understanding of how these materials interact with biological
systems might result in the creation of novel biomedical applications [2]. We present a study on the interaction of
model lipid membranes with gold nanoparticles (NP) of different surface modifications.
Neutron reflectometry experiments on zwitterionic DSPC and two-component negatively charged DSPC/DSPG
double bilayers were performed in the presence of gold nanoparticles (NP) functionalized with cationic and anionic
head groups. Structural information was obtained that provided insight into the fate of the NPs with regard to the
integrity of the model cell membranes.
The NPs functionalized with the cationic head groups penetrate into the hydrophobic moiety of the lipid bilayers and
cause membrane disruption at higher concentration. In contrast, the NPs functionalized with the anionic head
groups do not enter, but rather stabilize the lipid bilayer at alkaline pH [3].
The information obtained might influence the strategy for a better nanoparticle risk assessment based on a surface
charge evaluation and contribute to nano-safety considerations already during their design.
[1]
Maynard, A.D. Nanotechnology: a Strategy for Addressing Risk; Woodrow Wilson International Center for
Scholars: Washington, DC, (2006).
[2]
Murphy,C.J.; Gole, A.M.;Stone, J.W. Acc. Chem. Res. (2008), 41, 1721–1730.
[3]
Tatur, S.; Maccarini, M.; Barker, R.; Fragneto, G.; Langumir, submitted.
P.158 Investigation of liposome-based membranes formed by lipopolysaccharides and their interaction with hybrid
antimicrobial peptides
G Mangiapia1, R A Santagata1, G Vitiello1, G D'Errico1, A Silipo1, N Szekely2, and L Paduano1
1
Università degli Studi di Napoli , Italy, 2Jülich Centre for Neutron Science, Germany
Lipopolysaccharides (LPSs) are amphiphilic macromolecules indispensable for the growth and the survival of Gramnegative bacteria, one of the most diffuse classes of pathogens. They are composed of a hydrophilic
heteropolysaccharide unit, covalently linked to a lipophilic moiety called lipid A, which is embedded in the outer
leaflet and anchors these macromolecules to the lipid membrane. Here, we present a structural study on the
ICNS 2013 International Conference on Neutron Scattering
architecture and the conformation assumed by LPSs in the lipid membrane, along with the investigation of the
destabilizing effects caused by some hybrid antimicrobial peptides.
Investigations have been performed using an experimental strategy combining dynamic light scattering (DLS), small
angle neutron scattering (SANS), and electron paramagnetic resonance (EPR). These techniques are able to give
information on the liposome size and morphology, as well as on the dynamics of the bilayer.
Based on the collected data, LPSs aggregation behavior has been interpreted in terms of their complex molecular
architecture, and new insights on the liposome/peptide interaction process have been acquired. It has been shown
the first phase of the interaction process is characterized by the transition unilamellar → multilamellar liposomes
and it has also been proved that the process is strongly dependent on the presence of LPS molecules in the
liposome double layers.
P.159 Structure of polyunsaturated fatty acid lipid bilayers and the location of membrane guest molecules
D Marquardt1, F Heberle2, N Kucerka3, J Pan2, P Drazba2, R Standaert2, J Katsaras2 and T Harroun1
1
Brock University, Canada, 2Oak Ridge National Laboratory, USA, 3Canadian Neutron Beam Centre, Canada
Small angle neutron scattering (SANS) is a powerful technique for the study of biological molecules and systems.
Importantly, it allows for the structural determination of model membranes in their natural vesicle form in bulk
water. We measured SANS data for a variety of polyunsaturated phosphocholine lipids (PUFA-PC) with the following
sn1-sn2 acyl chain compositions: di-20:4, di-22:6, 16:0-20:4, 16:0-22:6, 18:0-20:4, and 18:0-22:6. For each
lipid, the fluid-phase temperature dependence of structural parameters was determined, followinga method for
calculating scattering density profiles (SDP) from the simultaneous analyses of X-ray and Neutron Scattering data
[Kučerka et al. BJ 95 (2008) 2356-2367]. This method then provides a more precise bilayer structure, including
area per lipid. As a result, a detailed characterization of PUFA-PCs will allow for the critical evaluation of the
umbrella interaction model for PC and cholesterol.
During this study we have also examined the prospect of determining the location of deuterium labeled small
molecules through the use of SANS. The prototypical neutron scattering method for the determination of a label
location in a model membrane is Bragg diffraction (SAND) from oriented (supported) multibilayer samples. We
intend to examine how label locations compare between the use of SANS and SAND, which will provide insight into
the quality of each experimental technique in terms of determining the location of small membrane bound
molecules. These results will then shed light on the mechanism of PUFA oxidation protection and its dependence on
the location of lipophilic antioxidant such as Vitamin E.
P.160 SANS investigation of membranes based on pervaporation
G Török1, L Vasilij2, K Yurii2 and K Svetlana3
1
Hungarian Academy of Sciences, Hungary, 2NRS Kurchatov Institute, Russia, 3Russian Academy of Sciences,
Russia
Polyamide-Imide pervaporation membranes (synthesized at IMC of RAS) dry and saturated with water have been
studied by small-angle neutron scattering to examine their structure as dependent on the conditions of formation.
There was shown that these membranes has a hard porous structure with sharp polymer borders. We have found
that the membranes’ structural elements (spherical voids, radii in the range 4-100 nm) are varying substantially as
defined by the time of formation, drying regimes, addition of fullerenes C60 , what treatment was performed to
modify mechanical and selective properties of membranes. The observed differences in the rates of membranes
swelling and its saturation level were interpreted as a consequence of existence of skin-layers at membranes
surface and the degree of the these surface regions hydrophilicity. The structural results were discussed in
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connection with the functional properties of these polymer systems to be used for a fine separation of azeoptropic
mixtures (e.g. water-ethanol).
P.161 Polymer induced swelling of solid supported lipid membranes studied by in-situ neutron reflectivity
M Trapp2, M Kreuzer1 and R Steitz3
1
Institut Catala de Nanotecnologia, Spain, 2University of Heidelberg, Germany, 3Helmholtz-Zentrum Berlin, Germany
Recently it has been found that oligolamellar DMPC lipid bilayer coatings do not detach from solid support against
excess aqueous phase at physiological temperature when incubated with hyaluronic acid (HA). [1] The coatings
rather adopt a highly ordered and stable new state as seen by specular and off-specular neutron reflectivity
measurements. That new state is best described as a hydrogel with a d-spacing approximately 380% larger than
that of the pure lipid coating. Latest neutron reflectivity experiments revealed that the observed swelling and
stabilization of the oligolamellar lipid linings is not unique to HA but is also invoked by synthetic polymers dissolved
in the excess aqueous solution. Negatively charged polymers (PSS) and positively charged polymers (PAH) both
interact with the solid supported zwitterionic DMPC membranes and both polymers induce a dramatic swelling of
the respective oligolamellar lipid coatings. On the base of resulting scattering length density profiles, we will discuss
a possible mechanism of the transformation of the oligolamellar lipid systems with incubation time, as well as
impacts of polymer charge and polymer size on the formation of the hydrogel layers. Also, potential consequences
of the new lamellar phase of the lipid linings with respect to natural processes like joint lubrication and wound
healing will be under consideration.
[1]
M. Kreuzer et al., Biochimica et Biophysica Acta - Biomembranes, 1818 , 11, 2012
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